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Novel insights into the Proteus mirabilis crystalline biofilm using real-time imaging

Novel insights into the Proteus mirabilis crystalline biofilm using real-time imaging
Novel insights into the Proteus mirabilis crystalline biofilm using real-time imaging
The long-term use of indwelling catheters results in a high risk from urinary tract infections (UTI) and blockage. Blockages often occur from crystalline deposits, formed as the pH rises due to the action of urease-producing bacteria; the most commonly found species being Proteus mirabilis. These crystalline biofilms have been found to develop on all catheter materials with P. mirabilis attaching to all surfaces and forming encrustations. Previous studies have mainly relied on electron microscopy to describe this process but there remains a lack of understanding into the stages of biofilm formation. Using an advanced light microscopy technique, episcopic differential interference contrast (EDIC) microscopy combined with epifluorescence (EF), we describe a non-destructive, non-contact, real-time imaging method used to track all stages of biofilm development from initial single cell attachment to complex crystalline biofilm formation. Using a simple six-well plate system, attachment of P. mirabilis (in artificial urine) to sections of silicone and hydrogel latex catheters was tracked over time (up to 24 days). Using EDIC and EF we show how initial attachment occurred in less than 1 h following exposure to P. mirabilis. This was rapidly followed by an accumulation of an additional material (indicated to be carbohydrate based using lectin staining) and the presence of highly elongated, motile cells. After 24 h exposure, a layer developed above this conditioning film and within 4 days the entire surface (of both catheter materials) was covered with diffuse crystalline deposits with defined crystals embedded. Using three-dimensional image reconstruction software, cells of P. mirabilis were seen covering the crystal surfaces. EDIC microscopy could resolve these four components of the complex crystalline biofilm and the close relationship between P. mirabilis and the crystals. This real-time imaging technique permits study of this complex biofilm development with no risk of artefacts due to sample manipulation. A full understanding of the stages and components involved in crystalline encrustation formation will aid in the development of new protocols to manage and ultimately prevent catheter blockage.
1932-6203
1-13
Wilks, Sandra A.
86c1f41a-12b3-451c-9245-b1a21775e993
Fader, Mandy J.
c318f942-2ddb-462a-9183-8b678faf7277
Keevil, C. William
cb7de0a7-ce33-4cfa-af52-07f99e5650eb
Wilks, Sandra A.
86c1f41a-12b3-451c-9245-b1a21775e993
Fader, Mandy J.
c318f942-2ddb-462a-9183-8b678faf7277
Keevil, C. William
cb7de0a7-ce33-4cfa-af52-07f99e5650eb

Wilks, Sandra A., Fader, Mandy J. and Keevil, C. William (2015) Novel insights into the Proteus mirabilis crystalline biofilm using real-time imaging. PLoS ONE, 10 (10), 1-13. (doi:10.1371/journal.pone.0141711).

Record type: Article

Abstract

The long-term use of indwelling catheters results in a high risk from urinary tract infections (UTI) and blockage. Blockages often occur from crystalline deposits, formed as the pH rises due to the action of urease-producing bacteria; the most commonly found species being Proteus mirabilis. These crystalline biofilms have been found to develop on all catheter materials with P. mirabilis attaching to all surfaces and forming encrustations. Previous studies have mainly relied on electron microscopy to describe this process but there remains a lack of understanding into the stages of biofilm formation. Using an advanced light microscopy technique, episcopic differential interference contrast (EDIC) microscopy combined with epifluorescence (EF), we describe a non-destructive, non-contact, real-time imaging method used to track all stages of biofilm development from initial single cell attachment to complex crystalline biofilm formation. Using a simple six-well plate system, attachment of P. mirabilis (in artificial urine) to sections of silicone and hydrogel latex catheters was tracked over time (up to 24 days). Using EDIC and EF we show how initial attachment occurred in less than 1 h following exposure to P. mirabilis. This was rapidly followed by an accumulation of an additional material (indicated to be carbohydrate based using lectin staining) and the presence of highly elongated, motile cells. After 24 h exposure, a layer developed above this conditioning film and within 4 days the entire surface (of both catheter materials) was covered with diffuse crystalline deposits with defined crystals embedded. Using three-dimensional image reconstruction software, cells of P. mirabilis were seen covering the crystal surfaces. EDIC microscopy could resolve these four components of the complex crystalline biofilm and the close relationship between P. mirabilis and the crystals. This real-time imaging technique permits study of this complex biofilm development with no risk of artefacts due to sample manipulation. A full understanding of the stages and components involved in crystalline encrustation formation will aid in the development of new protocols to manage and ultimately prevent catheter blockage.

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More information

Accepted/In Press date: 12 October 2015
e-pub ahead of print date: 30 October 2015
Published date: 30 October 2015
Organisations: Faculty of Health Sciences, Centre for Biological Sciences

Identifiers

Local EPrints ID: 385235
URI: http://eprints.soton.ac.uk/id/eprint/385235
ISSN: 1932-6203
PURE UUID: 7fa005d2-8ec0-46f9-be4d-fd6ec457cb18
ORCID for Sandra A. Wilks: ORCID iD orcid.org/0000-0002-4134-9415
ORCID for C. William Keevil: ORCID iD orcid.org/0000-0003-1917-7706

Catalogue record

Date deposited: 18 Jan 2016 13:39
Last modified: 15 Mar 2024 03:12

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